Abstract C51: Anticancer peptides PNC‐27/‐28: Cancer cell killers by binding to HDM2 and pore formation in cancer cell membranes

Studies in vitro and in vivo have clearly demonstrated the efficiency of the novel anti‐cancer peptides PNC‐27 and PNC‐28 in inducing the death of a variety of cancer cells by necrosis without any effect on normal cells. The peptides were derived from the HDM2‐binding domain of p53 (PNC‐27: AA12‐26; PNC‐28: AA17‐26) that were linked to membrane resident peptide (MRP) AA sequence from the homeodomain protein antennapedia of the fruit fly. Recently we have shown that in contrast to the necrotic cell death of cancer cells by external application of PNC‐28, the intracellular over‐expression of AAs17‐26 results in the cancer cells9 apoptosis while normal cells showed no response. We now provide evidence for the critical role for the expression of HDM2 in the plasma membrane of cancer cells and its absence in the plasma membrane of normal cells as a target molecule for PNC‐27. In direct competition experiments anti‐HDM2 antibodies (Ab) blocked PNC‐27 induced cell killing by >80% while control incubations with non‐specific Ab at identical protein concentrations did not hinder PNC‐27 from efficiently killing cancer cells. Immunofluorescence staining of HDM2 showed the presence of HDM2 on the plasma membrane of cancer cells (MIA‐PaCa‐2, BMRPA1.TUC3, A2058) which was absent in normal cell plasma membrane. Furthermore, co‐localization of HDM2 and PNC‐27 was observed on the plasma membrane of the cancer cells, suggesting their neighborliness. The findings were further confirmed by HDM2‐positive immunoblots (IB) of purified plasma membrane from several cancer cells while plasma membrane isolated from healthy normal cells only delivered negative IB results. Real‐time imaging using spinning disc confocal microscopy show that exposure to PNC‐27 of MIA PaCa‐2 cells preloaded with mitotracker dye in presence of propidium iodide (PI) results in the expansion within Citation Information: Cancer Res 2009;69(23 Suppl):C51.